DNA STRUCTURE AND DYNAMICS IN BAMHI-DNA INTERACTIONS

BAMHI-DNA 相互作用中的 DNA 结构和动力学

• 批准号: 6163485
• 负责人: Mary E Hatcher-Skeers
• 金额: $ 13.61万
• 依托单位: SCRIPPS COLLEGE
• 依托单位国家: 美国
• 财政年份: 2000
• 资助国家: 美国
• 起止时间: 2000-07-01 至 2003-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6163485
• 关键词: DNA; DNA binding protein; binding sites; chemical synthesis; computer simulation; deuterium; intermolecular interaction; molecular dynamics; nuclear magnetic resonance spectroscopy; nucleic acid sequence; nucleic acid structure; nucleotides; protein binding; structural biology

Recognition of specific DNA sequences by proteins plays an important role in many biological functions. Structural studies have shown that this specific recognition usually involves distortion of the DNA structure to accommodate protein binding. Two models for the mechanism of this specific recognition have emerged. In one, the protein plays an active role in recognition by rearranging itself and bending the DNA. It is suggested that the free energy released upon binding is sufficient to pay for that required for structural distortions. However, in order for complex formation to be favorable, the free energy required to bend the DNA and/or rearrange the protein must be low. Another model suggests that DNA possesses an intrinsic flexibility manifested in local, large amplitude dynamics. These flexible regions provide soft-spots which are easily distorted upon protein binding. In both these models, bending of the DNA must a low free energy process, suggesting that DNA has some inherent flexibility. In order to test this hypothesis, studies of local DNA dynamics must accompany structural investigations. In the next three years we plan to use high-resolution NMR studies and solid-state deuterium NMR studies to explore the local structural variations and dynamics of a DNA dodecamer containing the binding site for the BamHI restriction endonuclease. Specifically we propose the following program l) Synthesize deuterium labeled DNA phosphoramidites for incorporation into a DNA dodecamer containing the BamHI binding site. 2) Use solid-state deuterium NMR to characterize the local dynamics of backbone and sugar moieties of the individual nucleotides in the dodecamer. 3) Use high-resolution NMR to provide a structural description of the DNA dodecamer.

蛋白质对特定DNA序列的识别在许多生物学功能中发挥着重要作用。结构研究表明，这种特异性识别通常涉及 DNA 结构的扭曲以适应蛋白质结合。这种特定识别机制的两种模型已经出现。其中之一是，蛋白质通过自我重新排列和弯曲 DNA 在识别中发挥积极作用。这表明结合时释放的自由能足以支付结构扭曲所需的能量。然而，为了有利于复合物的形成，弯曲 DNA 和/或重新排列蛋白质所需的自由能必须较低。 另一种模型表明 DNA 具有内在的灵活性，这种灵活性体现在局部大振幅动力学上。这些柔性区域提供了在蛋白质结合时很容易变形的软点。在这两个模型中，DNA 的弯曲必须是一个低自由能过程，这表明 DNA 具有一定的固有灵活性。为了检验这一假设，局部 DNA 动力学研究必须与结构研究相结合。在接下来的三年中，我们计划使用高分辨率 NMR 研究和固态氘 NMR 研究来探索含有 BamHI 限制性内切核酸酶结合位点的 DNA 十二聚体的局部结构变化和动力学。具体来说，我们提出以下方案 l) 合成氘标记的 DNA 亚磷酰胺，用于掺入包含 BamHI 结合位点的 DNA 十二聚体中。 2) 使用固态氘核磁共振来表征十二聚体中各个核苷酸的主链和糖部分的局部动力学。 3) 使用高分辨率 NMR 提供 DNA 十二聚体的结构描述。

项目成果

The amide rotational barrier in isonicotinamide: Dynamic NMR and ab initio studies.

异烟酰胺中的酰胺旋转势垒：动态 NMR 和从头算研究。

• DOI: 10.1021/jp0460689
• 发表时间: 2005
• 期刊: The journal of physical chemistry. A
• 作者: Leskowitz,GarettM;Ghaderi,Nima;Olsen,RyanA;Pederson,Kari;Hatcher,MaryE;Mueller,LeonardJ
• 通讯作者: Mueller,LeonardJ